Neural precursor specification in the forebrain remains poorly understood, even though these precursors give rise to neurons that constitute essential circuitry for cognition, learning and memory. The initial specification of forebrain precursors is likely to rely upon signals from extrinsic sources that establish intrinsic organizing centers and regulate subsequent patterning and differentiation. The identity of such extrinsic sources, however, remains uncertain, and their influence on well-defined ventral and dorsal forebrain organizing centers is unknown. We have shown that inductive signaling between neural crest-derived frontonasal mesenchyme (FnM) and adjacent forebrain neuroepithelium influences early patterning, expression of signaling molecules, and subsequent precursor diversity in the forebrain. In Specific Aim 1 of this project we will determine whether this FnM-mediated induction is distinctly specified for forebrain patterning and facilitates regionally appropriate differentiation of forebrain neurons. It is unlikely that FnM is the sole source of extrinsic inductive signals for forebrain precursors. Our preliminary observations indicate that FnM acts in concert with another rarely considered source of signaling molecules available to the forebrain during early development: soluble proteins in the amniotic fluid (AF) when the anterior neural tube is open, and cerebrospinal fluid (CSF) once the anterior neural tube has closed. We have found that AF and CSF differentially support neurogenic capacity, regional identity, and subsequent proliferation of forebrain precursors. In Specific Aim 2, we will evaluate the activity of specific candidate signals in AF and CSF, and their effects-in concert with FnM-on neuroepithelial identity prior to anterior neural tube closure, as well as radial glial differentiation, and ventral and dorsal precursor distinctions once the neural tube closes. Finally, one molecule: retinoic acid (RA) has emerged as a candidate signal for essential aspects of FnM as well as AF/CSF influences on the developing forebrain. At early stages the FnM produces RA that acts on subsets of forebrain precursors. Subsequently, the meninges synthesize RA that acts on forebrain progenitors. In Specific Aim 3, we determine the specificity of these multiple RA sources for distinct steps in early forebrain patterning and precursor differentiation, as well as the consequences of initial RA signaling for forebrain precursor diversity and fate. Together, the experiments in our Specific Aims will define for the first time the specific contributions of extrinsic induction for establishing precursor, neuronal and circuit diversity in forebrain regions including the olfactory bulb, basal ganglia, and cerebral cortex. Our results will provide new insight into disrupted brain development in several disorders that also result in anomalous face, heart and limb development-all of which rely upon inductive mechanisms similar to that mediated by FnM-including autism, Down syndrome, mental retardation, and schizophrenia. PUBLIC HEALTH RELEVANCE: In behavioral disorders like autism, mental retardation, and psychiatric diseases like schizophrenia, abnormal brain function is thought to reflect disrupted early development of forebrain neurons and circuits. Understanding the role of signals that act on the developing forebrain, and how disrupting these signals changes the identity and fate of forebrain stem cells, is essential for defining pathogenic mechanisms in these increasingly prevalent diseases of forebrain development and function.